The present invention is related to a method for reducing the effects of mast cell mediated allergic reactions, including mast cell mediated allergy and asthma. In accordance with the present invention, these allergic reactions are reduced by administering a dehydroepiandrosterone (DHEA) derivative.
The publications and other materials used herein to illuminate the background of the invention, and in particular cases, to provide additional details respecting the practice, are incorporated by reference, and for convenience are numerically referenced in the following text and respectively grouped in the appended bibliography.
Dehydroepiandrosterone (DHEA), a weak androgen, serves as the primary precursor in the biosynthesis of both androgens and estrogens (1). DHEA has been reported to play a mitigating role in obesity, diabetes, carcinogenesis, autoimmunity, neurological loss of memory (2-5), and the negative effects of GCS on IL-2 production by murine T cells (6).
Recent insight into the mechanism of action of DHEA has come from studies of ischemia-induced reperfusion injury. The clinical term used to describe the pathological process of wound extension is progressive dermal ischemia and it appears to represent the consequences of a host-initiated, time-dependent reperfusion injury. DHEA, DHEAS, DHEA congeners and DHEA derivatives have been found to either reduce or protect thermally injured mice against reperfusion damage of the microvasculature. Additionally, intervention therapy with the active agent could be withheld for up to 4 hours after burn with substantial therapeutic benefit. It has been observed that the immediate response to a burn injury is in many ways similar to an experiment reperfusion injury in other tissues. Studies suggest that DHEA, either directly or indirectly, through its action on endothelium prevents damage to the microvasculature in reperfusion injury.
In another study the effect of DHEA on ischemia/reperfusion injury of the isolated rat cremaster muscle was evaluated. The experimental approach employed intravital microscopy to establish whether DHEA pre-treatment of rats prior to ischemia/reperfusion of the isolated muscle would protect against damage to the capillaries and venules of microcirculation. These studies indicated that in control animals, 6 hours of ischemia followed by re-flow analysis at 90 minutes and 24 hours lead to insufficient perfusion of the muscle. In DHEA pre-treated rats, 6 hours of ischemia followed by re-flow analysis at 90 minutes, 24 hours and even 4 days showed normal perfusion values in the isolated muscle. In addition, it was clear that the DHEA pre-treatment prevented sticking of neutrophils to endothelium. Additional studies in a global ischemic model demonstrated the protective effect of DHEA given intravenously after resuscitation of clinically dead rats.
Bacterial translocation is the process by which indigenous gut flora penetrate the intestinal barrier and invade sterile tissue. Included in this process is the migration of microbial organisms to the draining mesenteric lymph nodes, spleen, liver, blood and in some instances, the lung (7, 8). This phenomenon has been documented in humans following thermal injury (9-11) and ischemia-reperfusion injury (12). DHEA, DHEAS, DHEA congeners and DHEA derivatives have been found to either reduce or prevent bacterial translocation.
The evidence implicating the role of neutrophils in adult respiratory distress syndrome (ARDS) is substantial but indirect (13). Some of the first suggestions that neutrophils may cause an ARDS-like picture were found in severely neutropenic patients who were infused intravenously with donor neutrophils. Occasionally, within hours of neutrophil infusion, there was an abrupt "white-out" of the lungs (by x-ray) and onset of ARDS symptoms. Numerous studies have shown that neutrophils accumulate in the lung during ARDS. For example, their presence has been demonstrated histologically. During the early phases of ARDS, the number of circulating whole blood cells transiently decreases, probably due to their abnormal pulmonary sequestration. Some neutrophils that accumulate within lung capillaries leave the vascular space and migrate into the interstitium and alveolar airspaces. In normal healthy volunteers, neutrophils account for less than 3% of the cells that can be obtained by bronchoalveolar lavage (BAL). In patients with ARDS, the percentage of neutrophils in the lavage is markedly increased to 76-85%. The accumulation of neutrophils is associated with evidence of their activation. They demonstrate enhanced chemotaxis and generate abnormally high levels of oxygen metabolites following in vitro stimulation. Elevated concentrations of neutrophil secretory products, such as lactoferrin, have been detected in the plasma of patients with ARDS. Further evidence that neutrophils actively participate in lung injury was obtained from a clinical study of patients with mild lung injury who were neutropenic for an unrelated reason (e.g., receiving chemotherapy). It was noted that lung impairment frequently worsened if a patients hematological condition improved and circulating neutrophil counts recovered to normal levels.
As further proof that stimulated neutrophils can independently injure lung tissue, in vitro experiments have been performed using vascular endothelial and lung epithelial cells as targets. In some reports, neutrophils have been shown to detach endothelial cells or alveolar epithelial cells from the surface of the tissue culture dish. Obviously, if such an event were to occur in vivo, the denuded surfaces would permit substantial leakage of plasma contents. Furthermore, many reports have provided clear evidence that stimulated neutrophils are able to facilitate lysis of cultured vascular endothelial cells and alveolar epithelial cells. DHEA, DHEAS, DHEA congeners and DHEA derivatives have been found to either reduce or prevent ARDS.
In the United States, chronic obstructive pulmonary disease (COPD) represents the fifth most common cause of death (14). COPD also constitutes one of the most important causes of work incapacity and restricted activity (15). COPD, along with many other pulmonary diseases, causes pulmonary hypertension and right ventricular hypertrophy or cor pulmonale. Over 12 million patients in the United States alone have chronic bronchitis or emphysema, and approximately 3 million are chronically hypoxic with PaO.sub.2 &lt;60 mmHg. These patients develop hypoxic pulmonary vasoconstriction, and eventually, right ventricular hypertrophy (16). Once right ventricular hypertrophy develops, the three-year mortality rate of those patients is 60% (17, 18). Irrespective of the current management, morbidity and mortality of patients with COPD and pulmonary hypertension remain high.
One model to study pulmonary hypertension is the pulmonary vasoconstriction induced by alveolar hypoxia. Experiments in isolated animal (19) and human (20) pulmonary arteries suggest that hypoxia-induced pulmonary vasoconstriction is mediated by a direct effect of hypoxia on pulmonary vascular smooth muscle cell. It has been reported (21) that hypoxia can depolarize the pulmonary vascular smooth muscle membrane by inducing an increase in tissue Na.sup.+ and a decrease in K.sup.+. More recently, it has been reported that hypoxia can alter the membrane potential in rat main pulmonary artery smooth muscle cell and can stimulate Ca.sup.2+ influx through voltage-gated channels (22). There is strong evidence that Ca.sup.2+ entry blockade can attenuate hypoxic pulmonary vasoconstriction in isolated rat lung (23) and in patients with chronic obstructive lung disease (24). Conceivably, hypoxia may effect other membrane transport mechanisms that are involved in Ca.sup.2+ influx and/or efflux. For example, Voelkel et al. (25) speculated that hypoxia may impair Ca.sup.2+ extrusion. Farrukh et al. (26) has demonstrated that cAMP and cGMP reverse hypoxic pulmonary vasoconstriction by stimulating Ca.sup.2+ ATP-ase-dependent Ca.sup.2+ extrusion and/or redistribution. DHEA, DHEAS, DHEA congeners and DHEA derivatives have been found to either reduce or prevent pulmonary hypertension.
The above findings, as well as the finding that DHEA, DHEAS, DHEA congeners and DHEA derivatives reduce the expression of p-selectin by endothelial cells, are shown in, for example, U.S. Pat. Nos. 5,489,581; 5,532,230; 5,583,126; 5,587,369; and 5,635,496 and the published application of PCT/US95/10990, all incorporated by reference herein.
Allergic diseases are mediated, at least in part, by IgE antibody; IgE antibody production is a central feature of allergic diseases. These include food allergy, stinging insect allergy, latex allergy, and anaphylaxis, allergic rhinitis, and asthma. It will also deal briefly with diseases such as atopic dermatitis, whose pathogenesis is obscure but is likely to be related to other allergic diseases. The chapter focuses on human systems but includes some results with rodent models.
Allergic diseases affect 20% to 30% of the population of the United States (27). It may suggest some selective advantages to being a patient with these diseases. The majority of patients with allergic diseases are atopic. Atopic individuals produce IgE antibody to airborne allergens such as proteins in ragweed and/or grass pollens and/or dust mites, and they express allergic rhinitis and/or asthma and/or atopic dermatitis. Food allergy is often the first manifestation of allergic diseases in young atopic children. Moreover, there is a strong genetic component to the atopic state.
The expression of allergic disease requires a number of sequential events, including exposure to allergens, induction of IgE antibody production, binding of IgE to he surface receptors of mast cells and basophils, re-exposure to allergen, binding of allergen to cell-associated IgE, signal transduction in mast cells and basophils, mediator secretion, and mediator effects on end-organs such as blood vessels and bronchial smooth muscle.
As defined by Coombs and Gell (28), hypersensitivity reactions can be subdivided into four types, called 1,11, 111, and IV, which represent four distinct immune mechanisms that result in tissue injury. A subdivision of type IV reactions into IV A and IV B is also described below. This classification is outlined schematically in Table 1. These same four processes represent mechanisms of immune protection from infectious agents, as described below.
Type I reactions are "immediate hypersensitivity," or classical allergic reactions. These reactions occur within 15 mins following interaction of soluble antigen with mast cell-bound IgE antibody. The pathology is related to mast cell degranulation, and the reaction is driven by mast cell mediators such as histamine and leukotriene C4 (LTC4). An example of an in vivo counterpart is an urticarial reaction following injection of penicillin in a penicillin-allergic patient. The importance of type I reactions in protection from infectious organisms is uncertain, although the increased vascular permeability mediated by these reactions probably facilitates the capacity of antibody and inflammatory cells to arrive at the infected site (29).
Substances that induce symptoms of immediate hypersensitivity by inducing IgE antibody responses are termed allergens. Most atopic individuals produce IgE antibody to a long list of aeroallergens, that is, allergens found in the air. These allergens induce sensitization via exposure to the afferent immune system in the nasal or respiratory tract. A variety of allergens, derived from outdoor and indoor airborne sources, foods, and insect venoms, have been cloned and sequenced. The T-cell response pattern to allergens appears to be quite similar to that of conventional antigens, in that antigenic fragments are presented via MHC class II molecules on antigen-presenting cells to the T-cell receptor (30). Immunodominant peptides have been identified on several allergens; these have generally been DR-restricted, but recent studies have identified DP-restricted responses (31). The dose of exposure, the route of exposure (e.g., what type of particulate), and the genetic background of the host all interact to determine the magnitude of the IgE response to allergens. The levels of exposure to airborne allergens are quite low, suggesting that immune response genes may be identified that determine responsiveness to specific allergen epitopes (32). Moreover, the reasons why atopic patients produce IgE and make other immune responses to airborne allergens, while nonatopic patients do not, are not explained.
IgE antibodies are preferentially formed in response to parasitic antigens or allergens. Although low in concentration, IgE antibodies bind with high affinity to specific receptors (Fc.epsilon.RI) on mast cells and basophils. Antigen cross-linking of IgE molecules and the receptors to which they attach initiates the release or production of a variety of cellular mediators. The mediators begin a series of physiologic events that lead to allergic diseases, such as allergic rhinitis, asthma and urticaria, but they may also help to confer specific protective immunity against parasites.
Antigen-mediated crosslinking of Fc.epsilon.RI results in secretion of mediators from mast cells. Both the morphology of the mast cells and the mediator levels in tissue fluids confirm that mast cell degranulation occurs in vivo during allergic reactions (33, 34). The mediators secreted by mast cells and basophils account for the symptoms of allergic reactions (35). These include the following preformed mediators, which are associated with granules: histamine (bound to sulfated proteoglycans, either heparin or chondroitin sulfate), the proteoglycans themselves, and several proteases, including the neutral proteases, carboxypeptidase(s), tryptase, and (in some mast cells) chymase. The cytokine TNF-.alpha. is released in part from a stored form in mast cells (36), but this cytokine is not stored in macrophages or T cells. In addition, there are newly synthesized molecules, including LTC4, PGD2, and PAF, and cytokines.
Asthma is a chronic disease of the large and small airways of the lung (37-39) which affects 5% to 10% of the population. The disease is more common in children, but may persist for years and may develop only in adult life. Asthma is characterized by several clinical and pathological features. The most prominent feature is bronchospasm, or narrowing of the airways; the bronchospasm is often reversible over time or with treatment. Asthmatic patients have prominent contraction of the smooth muscle of large and small airways, an increased mucus production, and an inflammatory infiltrate consisting of eosinophils, as well as basophils and T lymphocytes; epithelial cell shedding occurs (40, 41). Airway narrowing is due not only to bronchial smooth muscle contraction, but also to mucus production and inflammation. Important laboratory findings include evidence of airway narrowing, increased numbers of circulating eosinophils, and moderate increases in total serum IgE (compared to nonasthmatic patients of the same age). A substantial number of patients are atopic and a substantial number express IgE antibody against specific allergens such as dust mite (42). One additional finding is airway hyperreactivity. That is, while stimuli that induce smooth muscle contraction, such as histamine and methacholine (an acetylcholine-like agent), may induce bronchospasm in all individuals, much lower concentrations of these bronchospastic agents are required in order to induce bronchoconstriction in hyperreactive individuals.
The mechanisms that induce all the pathologic findings in asthma are not known. In many patients with asthma, allergen exposure may induce a fall-blown, severe episode of airway inflammation. In such patients, the mechanisms are presumed to be the same as those that induce a pulmonary LPR after inhalation of allergen: that is, allergen crosslinks mast cell-associated IgE antibody, which in turn leads to the release of mast cell mediators.
Mast cell mediators such as histamine and LTC4 are important inducers of bronchospasm and mucus production. Cytokines, perhaps derived from mast cells or T cells (which may interact with antigen processed by an antigen-presenting cell), induce inflammation. Eosinophil-derived mediators, such as major basic protein, peroxidase, and cationic protein, appear to be important in inducing epithelial injury (40, 43). There also appear to be antigen-independent mechanisms of inducing asthma, including viral infection and exercise. It is possible that these other mechanisms are also initiated by a common pathway of mast cell activation (although many investigators believe that mast cells are not of central importance). It is likely that eosinophils are an important mediator of asthma; in patients, levels of circulating eosinophils increase when asthma worsens. Moreover, glucocorticoids are effective in treating moderate and severe asthma and in reducing levels of circulating and tissue eosinophils.
Several abnormalities may be present in patients with asthma. They not only tend to be atopic, and thus have increased tendency to produce IgE antibodies to allergens, but their basophils tend to secrete mediators more readily in response to certain stimuli (44). Furthermore, many patients with asthma have been reported to have several abnormalities of autocrine or neuropeptide receptors. Some years ago, it was noted that asthmatic patients had a generalized decrease in .beta.-adrenergic receptor (which mediates smooth muscle relaxation) responsiveness and increased cholinergic and .beta.-adrenergic (which mediates smooth muscle contraction) responsiveness; indeed, some of these patients have circulating antibodies to .beta.-adrenergic receptors. However, these findings are not specific for asthma (45). More recently, asthmatic patients have been reported to have a decrease in receptors for vasoactive intestinal peptide (a ligand that relaxes smooth muscle) and perhaps an increase in receptors for substance P (a ligand that contracts smooth muscle) (46, 47).
Several non-IgE pathways result in asthma. Viral infections are associated with concomitant worsening of pulmonary function (38). Nonsteroidal anti-inflammatory agents, such as aspirin, may exacerbate asthma; about 5% of asthmatic patients are sensitive to these agents (48). It has been hypothesized that these agents act by altering the metabolism of arachidonic acid, since these agents block prostaglandin synthetase. However, the precise mechanism is unknown. Another cause of asthma is exercise, apparently because of a fall in temperature and humidity of the airway. The mechanisms are not clear. One interesting hypothesis, that exercise-induced asthma results from inducing local hyperosmolarity, which is in turn a trigger for mast cell activation, has not been confirmed. Among all these non-IgE pathways, whether mast cell mediator release has a role is arguable.
One of the most interesting areas of recent investigation concerns the role of allergens. Case-control studies of emergency room admissions with asthma have established that IgE antibodies to certain allergens, namely, "indoor allergens" from dust mite, cat, and cockroach, are important risk factors (37, 49). Other studies have shown that, in allergic patients, inhalational challenge with these allergens induces an inflammatory LPR and bronchial hyperreactivity. Dust mite-sensitive patients with asthma, when moved into an environment free of dust mites, may exhibit a dramatic improvement in symptoms (50). These latter findings are provocative, although they need to be repeated in a controlled study. Unexpectedly, improvement of some patients required months; there is no obvious explanation for this. As previously noted, other studies with dust mites suggest that high dust mite exposure in the first two years of life is predictive of the presence of asthma at age 10. Consequently, environmental control of these allergens is being tested for its effectiveness in treating asthma. In addition, immunotherapy (see below) is effective in treating some patients with allergen-induced asthma.
Environmental factors other than allergens may be important in asthma. Certain chemicals, such as ozone and nitric oxide, are reported to worsen asthma (51, 52). Also, passive cigarette smoke exposure worsens asthma (53).
Within the last 10 years, the incidence of asthma, its severity, and deaths from asthma have increased. The increase in asthma morbidity and mortality is most striking in children, and in the United States the morbidity and mortality are highest in African-American children in the inner city (54). These epidemiologic trends have not yet been adequately explained. One interesting idea is that, in attempts to improve the energy efficiency of homes, these homes have become "tighter" and less leaky and have allowed the concentrations of allergens and other adverse environmental factors to increase (55).
It is desired to identify compounds which are useful in the treatment of mast cell mediated allergic reactions, including type I hypersensitivity response to allergens and asthma.